Oral Presentation Australasian RNA Biology and Biotechnology Association 2024 Conference

Therapeutic efficacy of antisense oligonucleotides in humanized preclinical mouse model of inherited childhood-lethal neurodegenerative disorder (108574)

Raman Sharma 1 , Mark Corbett 1 , Rudrarup Bhattacharjee 1 2 , Tarin Ritchie 1 , Shreya Agarwala 1 , Nicholas JC Smith 3 , David A Stroud 4 , Daniella Hock 4 , Kim M Hemsley 5 , Melissa White 6 , Paul Thomas 6 , Jozef Gecz 1
  1. Adelaide Medical School and the Robinson Research Institute, The University of Adelaide, Adelaide, SA5000, Australia
  2. Adelaide Centre for Epigenetics, School of Biomedicine, The University of Adelaide, Adelaide, SA, Australia
  3. Department of Neurology, Women’s and Children’s Health Network, Adelaide, SA, Australia
  4. Department of Biochemistry and Pharmacology, Bio21 Molecular Science and Biotechnology Institute, University of Melbourne, Parkville, Victoria, Australia
  5. Childhood Dementia Research Group, College of Medicine and Public Health, Flinders Health & Medical Research Institute, Flinders University, Bedford Park, South Australia, Australia
  6. South Australian Genome Editing Facility, SAHMRI and The Robinson Research Institute, The University of Adelaide, Adelaide, SA, Australia

TIMMDC1 encodes the Translocase of Inner Mitochondrial Membrane Domain-Containing protein 1 subunit of complex I of the electron transport chain responsible for ATP production. We reported complete restoration of TIMMDC1 mRNA, protein, complex I proteome and mitochondrial function in cells of patients with a pathogenic TIMMDC1 c.596+2146A>G,sing two different Splice-Switching Oligonucleotides (SSOs) (Kumar et al Npj Genom Med 7:9, 2022). More than 13 patients with a severe early-onset, progressive, and fatal neurodegenerative disorder have been reported to carry this ancestral TIMMDC1 variant globally so far. For assessing SSO efficacy for eventual patient treatment, we have generated a humanized Timmdc1 c.597-1340A>G mouse model by inserting the human variant-containing intron sequence into the orthologous mouse Timmdc1 gene. The homozygous mutant mice recapitulate human phenotype e.g., severe tremors and righting problems (an indication of motor neuron degeneration) from post-natal day P7 and death by P12. RNA analyses of Timmdc1 mutant mouse fibroblasts and tissues (brain, quadriceps, liver) showed poison exon inclusion into the mouse Timmdc1 mRNA in a manner identical to human patient fibroblasts. This aberrant Timmdc1 mRNA is degraded via non-sense mediated mRNA decay. Quantitative proteome (heart, kidney, liver and brain) and western blot (brain, liver, quadriceps and eye) analyses show significantly reduced TIMMDC1 protein in Timmdc1 homozygous mutant compared to the wild type mice. We are currently testing therapeutic efficacy of SSOs by delivering centrally (intracerebroventricular, ICV) to increase Timmdc1 expression in central nervous system (CNS), systemically (subcutaneous, SC) to increase Timmdc1 expression in different tissues or combined (ICV and SC) to homozygous mutant and their wild type/heterozygous littermate pups. The results will expedite translation to clinical trials with a view to providing potentially life-saving treatment to individuals with mitochondrial disease caused by the TIMMDC1 c.596+2146A>G variant (in a homozygous or compound heterozygous state).